Osteoporosis is a disease characterized by low bone mineral density (BMD), the strongest predictor of fracture risk. It is now well known that BMD variance is determined by both genetic and environmental factors. Accordingly, studies in human and animal models have revealed evidence for the presence of several quantitative trait loci (QTL) that contribute to BMD variations in humans and experimental animals. However, the identification of BMD QTL genes remains a big challenge. In our recent study, we have identified duffy antigen receptor for chemokines (Darc) as a BMD QTL gene in chromosome 1 which regulates BMD negatively by increasing osteoclast differentiation. Darc binds to chemokines known to be involved in osteoclast differentiation. In terms of mechanism by which Darc regulates osteoclasts, our preliminary studies have shown that 1) M-CSF dependent bone marrow cells derived from Darc-knockout (Darc-KO) mice exhibit reduced binding to chemokines such as MCP-1 and RANTES;2) femurs isolated from Darc-KO mice showed reduced TRAP positive bone resorbing surface compared to control mice. In contrast, bone formation parameters were similar in both lines of mice;and 3) mouse monocyte RAW264.6 cells overexpressing Darc exhibited significant increase in transendothelial migration (TEM) towards the chemokine RANTES compared to RAW cells transfected with empty vector. These data demonstrate that osteoclast TEM towards chemokines is in part regulated by Darc. Based on these and other preliminary data that Darc-KO mice exhibit higher BMD compared to wild type mice and the known action of Darc in neutrophils/monocyteTEM, we propose the hypothesis that Darc regulates the migration of osteoclast precursor (OCP) cells through chemokines for subsequent fusion and differentiation. To test this hypothesis, we will 1) determine if the overexpression and the blockage of Darc expression in endothelial cells or in OCP cells will affect the transmigration of OCP cells towards MCP-1 and RANTES chemokines;2) induce chemokine expression in Darc-KO, transgenic mice that overexpress Darc only in endothelial cells and control mice by treatment with lipopolysaccharide and evaluate the consequence of increased chemokine expression on OCP recruitment and bone resorption in vivo. We anticipate that successful completion of this study would lead to increased understanding of the role of Darc-chemokine interaction in regulating osteoclast development and inflammation-induced bone loss. The confirmation of our hypothesis that Darc regulates osteoclast activity via binding to chemokines could lead to development of diagnostic screens to identify patients who are at increased risk for osteoporosis and therapeutic strategies to treat osteoporotic patients with chemokine-induced high bone turnover. PUBLIC HEALTH RELEVANCE: Low peak bone mineral density (BMD) has been established as an important risk factor for osteoporosis. Because >70% variation in peak BMD is genetically determined, it is of great importance to identify the genes and their mechanisms that contribute to variation in peak BMD. In this regard, we have recently identified that genetic variation in duffy antigen receptor for chemokines (Darc) regulates BMD negatively by influencing osteoclastic bone resorption. This study will test a novel hypothesis that DARC regulates osteoclast mobility by modulating the activity of chemokines. Our confirmation of the hypothesis that Darc regulates osteoclast function by binding to chemokines could lead to future development of diagnostic screens to identify patients who are at risk for osteoporosis and therapeutic strategies to treat osteoporotic patients with chemokine- induced high bone turnover.